Assays for the detection of target species such as cholera toxin and Bacillus anthracis protective antigen (PA) have been known using phospholipids bilayer membranes supported on a silica-coated waveguide platform (see, Martinez et al., J. Mat. Chem., vol. 15, pp. 4639-4647, (2005)). While lipid membranes offered excellent resistance to non-specific binding, lipid membranes are not robust and do not endure either prolonged storage or use under harsh conditions.
In the past, many groups have explored various PEGyolated SAMs that have a short attachment group to the oxide surface with terminal polyethylene glycols (PEGs) of varying lengths. Often, the synthetic route to these earlier SAMs was through use of either a methyl-diethoxy-silane or a methyl-dimethoxy-silane. Although previous SAMs based on this approach showed good antifouling properties (non-specific binding) using optical microscopy, they did not exhibit good non-specific binding when using a waveguide-based sandwich assay approach. The reason for this difference is the relatively high optical intensity at the surface of the planar optical waveguide relative to the optical field intensity used in confocal microscopy. Essentially, the use of evanescent excitation is much more sensitive.
The failures of earlier diethoxy-methyl-aminopropylsilane-based SAMs in waveguide assays prompted reevaluation of surface chemistries. Whitesides and Grunze provide examples of PEG-terminated alkylthiols that are very good at resisting non-specific protein adsorption when analyzed by fluorescence microscopy (see Grosdemange et al., J. Am. Chem. Soc., vol. 113, pp. 13-20 (1993) and Herrwerth et al., J. Am. Chem. Soc., vol. 125, pp. 9359-9366 (1993)). The advantages of their films include dense packing due to the hydrophobic interactions of the alkyl chains, as well as the hydrophilicity of the terminal polyethylene glycol units. However, their films are prepared on metal surfaces, e.g., silver and gold. These films would not work for oxide surfaces like those used in the currently desired optical methods.